We have prepared large quantities of highly purified tetanus toxin by both the culture filtrate and cell extract procedure. We have separated the toxin molecule into its heavy chain (100,000 daltons) and its light chain (50,000 daltons) components by preparative polyacrylamide gel electrophoresis. By the omission or addition of reducing agents we are able to prepare two species each of both the H and L chain. We are also able to prepare both nicked (extracellular) and unnicked (intracellular) toxin in highly purified form. The amino and carboxy terminal residues of both toxins and the major peptides are being determined. We chose the L chain to start our sequence efforts and have determined some 22 residues from the amino terminal. We have also treated the L chain with cyanogen bromide and have fragmented it into at least eight peptides which are separated by SDS-urea polyacrylamide gels. Amino terminal analysis of the cleavage mixture revealed eight amino terminals in approximately equal molar quantities. We are currently separating these peptides and will sequence each one as we isolate it. We are continuing these efforts and hope to complete as near as possible the complete amino acid sequence of the L chain this year.